PROJECT SUMMARY A prominent feature of multiple myeloma (MM) and other malignancies is significant genomic instability leading to clonal evolution and disease progression. We previously reported that homologous recombination (HR), the error free DNA repair system in normal cellular environment, is dysregulated in MM as well as esophageal cancer, and contributes to genomic instability, development of drug resistance, and tumor growth. In the previous funding period, we observed significantly elevated nuclease activity in newly-diagnosed and relapsed MM compared to MGUS patient samples. A functional siRNA screen identified that two members of apurinic/apyrimidinic (AP) nuclease activity (APEX 1 and 2) are amongst the most prominent contributors to dysregulated nucleolytic and HR activities. We also observed that the high expression of APEX1 and APEX2 correlated with increased copy number events and poor survival in MM. In our preliminary studies we observe that transgenic and chemical inhibition of AP nuclease activity inhibited DNA breaks, HR activity, and genomic instability in MM cells, as well as induced a strong G2/M arrest; while transgenic upregulation of AP activity increased DNA breaks, HR activity, genomic instability, and led to oncogenic transformation in normal human cells as well as tumorigenesis in zebrafish and mouse models. We, therefore, hypothesize that elevated AP expression drives genomic instability and clonal evolution, and may represent a potential therapeutic target. To investigate the role of APEX nucleases in MM and to evaluate their therapeutic potential, we will To investigate the role of APEX nucleases (APEX1 and 2) in genomic evolution in MM (Sp. Aim 1); investigate APEX1/APEX2- induced genomic instability and malignant transformation in a transgenic mouse model of B cell malignancy (Aim 2); and evaluate inhibitors of AP nuclease activity in MM (Aim 3) alone and in combination with other anti-MM agents in vitro, in vivo and in Phase I/II clinical study in relapsed refractory multiple myeloma. This project will help identify novel target as well as drugs that may help inhibit tumor growth, prevent/delay genomic evolution, which in turn may help make MM cells genomically stable.